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Transforming growth factor ? (TGF?) cross-talk with the unfolded protein response is critical for hepatic stellate cell activation.

ABSTRACT: Transforming growth factor ? (TGF?) potently activates hepatic stellate cells (HSCs), which promotes production and secretion of extracellular matrix (ECM) proteins and hepatic fibrogenesis. Increased ECM synthesis and secretion in response to TGF? is associated with endoplasmic reticulum (ER) stress and the unfolded protein response (UPR). TGF? and UPR signaling pathways are tightly intertwined during HSC activation, but the regulatory mechanism that connects these two pathways is poorly understood. Here, we found that TGF? treatment of immortalized HSCs (i.e. LX-2 cells) induces phosphorylation of the UPR sensor inositol-requiring enzyme 1? (IRE1?) in a SMAD2/3-procollagen I-dependent manner. We further show that IRE1? mediates HSC activation downstream of TGF? and that its role depends on activation of a signaling cascade involving apoptosis signaling kinase 1 (ASK1) and c-Jun N-terminal kinase (JNK). ASK1-JNK signaling promoted phosphorylation of the UPR-associated transcription factor CCAAT/enhancer binding protein ? (C/EBP?), which is crucial for TGF?- or IRE1?-mediated LX-2 activation. Pharmacological inhibition of C/EBP? expression with the antiviral drug adefovir dipivoxil attenuated TGF?-mediated activation of LX-2 or primary rat HSCs in vitro and hepatic fibrogenesis in vivo Finally, we identified a critical relationship between C/EBP? and the transcriptional regulator p300 during HSC activation. p300 knockdown disrupted TGF?- or UPR-induced HSC activation, and pharmacological inhibition of the C/EBP?-p300 complex decreased TGF?-induced HSC activation. These results indicate that TGF?-induced IRE1? signaling is critical for HSC activation through a C/EBP?-p300-dependent mechanism and suggest C/EBP? as a druggable target for managing fibrosis.

SUBMITTER: Liu Z

PROVIDER: S-EPMC6398135 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

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Transforming growth factor β (TGFβ) cross-talk with the unfolded protein response is critical for hepatic stellate cell activation.

Liu Zhikui Z Li Chao C Kang Ningling N Malhi Harmeet H Shah Vijay H VH Maiers Jessica L JL

The Journal of biological chemistry 20190104 9

Transforming growth factor β (TGFβ) potently activates hepatic stellate cells (HSCs), which promotes production and secretion of extracellular matrix (ECM) proteins and hepatic fibrogenesis. Increased ECM synthesis and secretion in response to TGFβ is associated with endoplasmic reticulum (ER) stress and the unfolded protein response (UPR). TGFβ and UPR signaling pathways are tightly intertwined during HSC activation, but the regulatory mechanism that connects these two pathways is poorly unders ...[more]

PMID: 30610118

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Project description:Hepatic stellate cells activate upon liver injury and help at restoring damaged tissue by producing extracellular matrix proteins. A drastic increase in matrix proteins results in liver fibrosis and we hypothesize that this sudden increase leads to accumulation of proteins in the endoplasmic reticulum and its compensatory mechanism, the unfolded protein response. We indeed observe a very early, but transient induction of unfolded protein response genes during activation of primary mouse hepatic stellate cells in vitro and in vivo, prior to induction of classical stellate cell activation genes. This unfolded protein response does not seem sufficient to drive stellate cell activation on its own, as chemical induction of endoplasmic reticulum stress with tunicamycin in 3D cultured, quiescent stellate cells is not able to induce stellate cell activation. Inhibition of Jnk is important for the transduction of the unfolded protein response. Stellate cells isolated from Jnk knockout mice do not activate as much as their wild-type counterparts and do not have an induced expression of unfolded protein response genes. A timely termination of the unfolded protein response is essential to prevent endoplasmic reticulum stress-related apoptosis. A pathway known to be involved in this termination is the non-sense-mediated decay pathway. Non-sense-mediated decay inhibitors influence the unfolded protein response at early time points during stellate cell activation. Our data suggest that UPR in HSCs is differentially regulated between acute and chronic stages of the activation process. In conclusion, our data demonstrates that the unfolded protein response is a JNK1-dependent early event during hepatic stellate cell activation, which is counteracted by non-sense-mediated decay and is not sufficient to drive the stellate cell activation process. Therapeutic strategies based on UPR or NMD modulation might interfere with fibrosis, but will remain challenging because of the feedback mechanisms between the stress pathways.

Project description:Background & aimsActivation of Kupffer cells and recruitment of monocytes are key events in fibrogenesis. These cells release soluble mediators which induce the activation of hepatic stellate cells (HSCs), the main fibrogenic cell type within the liver. Mer tyrosine kinase (MerTK) signaling regulates multiple processes in macrophages and has been implicated in the pathogenesis of non-alcoholic steatohepatitis-related fibrosis. In this study, we explored if MerTK activation in macrophages influences the profibrogenic phenotype of HSCs.MethodsMacrophages were derived from THP-1 cells or differentiated from peripheral blood monocytes towards MerTK+/CD206+/CD163+/CD209- macrophages. The role of MerTK was assessed by pharmacologic and genetic inhibition. HSC migration was determined in Boyden chambers, viability was measured by the MTT assay, and proliferation was evaluated by the BrdU incorporation assay.ResultsGas-6 induced MerTK phosphorylation and Akt activation in macrophages, and these effects were inhibited by UNC569. During polarization, MerTK+/CD206+/CD163+/CD209- macrophages exhibited activation of STAT3, ERK1/2, p38 and increased expression of VEGF-A. Activation of MerTK in THP-1 macrophages induced a secretome which promoted a significant increase in migration, proliferation, viability and expression of profibrogenic factors in HSCs. Similarly, conditioned medium from MerTK+ macrophages induced a significant increase in cell migration, proliferation, STAT3 and p38 phosphorylation and upregulation of IL-8 expression in HSCs. Moreover, conditioned medium from Gas-6-stimulated Kupffer cells induced a significant increase in HSC proliferation. These effects were specifically related to MerTK expression and activity in macrophages, as indicated by pharmacologic inhibition and knockdown experiments.ConclusionsMerTK activation in macrophages modifies the secretome to promote profibrogenic features in HSCs, implicating this receptor in the pathogenesis of hepatic fibrosis.Lay summaryFibrosis represents the process of scarring occurring in patients with chronic liver diseases. This process depends on production of scar tissue components by a specific cell type, named hepatic stellate cells, and is regulated by interaction with other cells. Herein, we show that activation of MerTK, a receptor present in a population of macrophages, causes the production of factors that act on hepatic stellate cells, increasing their ability to produce scar tissue.

Dataset Information

Transforming growth factor ? (TGF?) cross-talk with the unfolded protein response is critical for hepatic stellate cell activation.

Publications

Transforming growth factor β (TGFβ) cross-talk with the unfolded protein response is critical for hepatic stellate cell activation.

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